Stem cells have significant therapeutic implications and importance in the treatment of disease and pathological conditions in humans and other animals. Development of stem cell lines that produce or regenerate tissues of the human body has the potential to radically improve the quality and length of life. As an example, stem cells present opportunities to overcome tissue rejection by a patient's immune system. Stem cells might also be modified through genetic changes or other techniques to overcome immune rejection and/or to act as a vehicle for in situ delivery of other therapeutic agents.
In certain clinical therapies, contact with foreign or undefined proteins should be avoided in order to enhance stem cell use in ex vivo preparation of cells, genes and tissue therapies. In addition, isolation of a progenitor/stem cell enriched population would enhance predictability of fabrication and long-term graft success of tissue engineered constructs.
Tissue engineering and regenerative medicine has created considerable interest in the clinical application of stem cells to both regenerate body tissues and to deliver genetic material. As only one example among others, the production of a specific gene-transduced oral mucosal graft that can be used for reconstruction of major oral defects would be an asset to reconstructive surgery and/or gene therapy. The graft would then act both as a material for reconstruction and as a repository for in situ transmucosal delivery of recombinant growth factors or cytokines. Primary human keratinocytes fulfill most requirements for use in these treatment modalities. Compared with other types of human cells, keratinocytes can grow to generate cohesive sheets of epithelium for grafting onto patients. In addition, the transplanted grafts are readily visible over time allowing constant control and follow up as well as the local modulation of transgene expression using an appropriate promoter (Serrano et al. 2003). Long term survival of stably transduced epidermal cells in a graft can allow the secretion of exogenous gene products into the bloodstream (Cao et al., 2002, Pfützner et al., 2002, Rollman et al., 2003).
Several barriers presently exist that have impeded this technology from moving into the clinical arena. First is the ability to fabricate these “smart” grafts in a more efficient and robust manner, i.e. to develop a more highly proliferative and expanding cell population. Second is the ability to isolate a stem/progenitor cell, thus making gene therapy more practical by achieving high-level gene expression in a significant percentage of cells. Lastly, if stem cells are to play a role in clinical therapies the cells should not come into contact with foreign or undefined proteins (animal serum or feeder layer cells, or pituitary extract) in order to gain FDA acceptance.
The ability to isolate an epidermal stem cell for use in fabrication of autologous grafts would result in a more robust engineered construct capable of a higher level of gene expression in a significant percentage of cells, thus creating a more predictable graft success and functionality for long term use as a vehicle for gene delivery (Bianco and Robey, 2001, Ortiz-Urda et al., 2002, Chen M et al., 2002). The development of an ex vivo produced oral mucosa equivalent, using an oral keratinocyte progenitor/stem cell-enriched population, with long-term in vitro growth properties, that persist in vivo and forms a fully differentiated epidermis (Kolodka et al., 1998), would allow the generation of a more robust and functional EVPOME that has a higher proliferative capacity and longer life-span. To our knowledge, the identification, isolation and fabrication of human oral mucosal grafts with human oral keratinocyte stem cells has not occurred.
Substantial scientific and ethical challenges remain. In addition, much controversy remains over the sources of biological materials for stem cell research. Thus an unmet need remains for additional sources of stem cells that avoid or mitigate these and other issues.